Toxic Effect of Metal Doping on Diatoms as Probed by Broadband Terahertz Time-Domain Spectroscopy.

The global marine environment is increasingly affected by human activities causing climate change, eutrophication, and pollution. These factors influence the metabolic mechanisms of phytoplankton species, such as diatoms. Among other pollutant agents, heavy metals can have dramatic effects on diatom viability. Detailed knowledge of the interaction of diatoms with metals is essential from both a fundamental and applicative point of view. To this aim, we assess terahertz time-domain spectroscopy as a tool for sensing the diatoms in aqueous systems which mimic their natural environment. Despite the strong absorption of terahertz radiation in water, we show that diatoms can be sensed by probing the water absorption enhancement in the terahertz range caused by the water-diatom interaction. We reveal that the addition of metal dopants affects this absorption enhancement, thus enabling the monitoring of the toxic effects of metals on diatoms using terahertz spectroscopy. We demonstrate that this technique can detect the detrimental effects of heavy metals earlier than conventional methods such as microscopy, enzymatic assays, and molecular analyses aimed at assessing the overexpression of genes involved in the heavy metal-stress response.

Long-term holographic phase-contrast time lapse reveals cytoplasmic circulation in dehydrating plant cells.

The intracellular dynamics of onion epidermal cells during the dehydration process is observed by holographic microscopy. Both the nucleus and cytoplasm are accurately revealed by quantitative phase imaging while dehydration takes place. Indeed, we notice that the contrast of phase images increases with the decrease in cellular water content. We foresee that such a dehydrating process can be effective for improving phase contrast, thus permitting better imaging of plant cells with the scope of learning more about cellular dynamics and related phenomena. Exploiting this concept, we observe intracellular cytoplasmic circulation and transport of biological material.

On-speckle suppression in IR digital holography.

Long-IR wavelength is the best option for capturing digital holograms of large-size, real-world objects. However, the coherent noise level in a long-IR hologram is by far larger than that of a visible wavelength recording, thus resulting in a poor quality of both numerical and optical reconstructions. In this Letter, we show how such coherent noise can be efficiently suppressed by employing an optical scanning multi-look approach, in combination with 3D block matching numerical filtering. Results demonstrate the possibility to obtain near noise-free numerical reconstructions of IR digital holograms of large-size objects, while preserving resolution. We applied this method to the holograms of a rotating statuette. It will be shown that a remarkable contrast enhancement is achievable along with the recovery of object details that otherwise would be lost because of large speckle grains intrinsically due to the source coherence.

Encoding multiple holograms for speckle-noise reduction in optical display.

In digital holography (DH) a mixture of speckle and incoherent additive noise, which appears in numerical as well as in optical reconstruction, typically degrades the information of the object wavefront. Several methods have been proposed in order to suppress the noise contributions during recording or even during the reconstruction steps. Many of them are based on the incoherent combination of multiple holographic reconstructions achieving remarkable improvement, but only in the numerical reconstruction i.e. visualization on a pc monitor. So far, it has not been shown the direct synthesis of a digital hologram which provides the denoised optical reconstruction. Here, we propose a new effective method for encoding in a single complex wavefront the contribution of multiple incoherent reconstructions, thus allowing to obtain a single synthetic digital hologram that show significant speckle-reduction when optically projected by a Spatial Light Modulator (SLM).

Common-path configuration in total internal reflection digital holography microscopy.

Total Internal Reflection Digital Holographic Microscopy (TIRDHM) is recognized to be a powerful tool for retrieving quantitative phase images of cell-substrate interfaces, adhesions, and tissue structures close to the prism surface. In this Letter, we develop an improved TIRDHM system, taking advantage of a refractive index mismatch between the prism and the sample substrate, to allow phase-shifting DH with just a single-beam interferometric configuration. Instead of the traditional off-axis method, phase-shift method is used to retrieve amplitude and phase images in coherent light and TIR modality. Essentially, the substrate-prism interface acts like a beam splitter generating a reference beam, where the phase-shift dependence on the incident angle is exploited in this common-path configuration. With the aim to demonstrate the technique's validity, some experiments are performed to establish the advantage of this compact and simple configuration, in which the reference arm in the setup is avoided.

Refocusing criterion via sparsity measurements in digital holography.

Several automatic approaches have been proposed in the past to compute the refocus distance in digital holography (DH). However most of them are based on a maximization or minimization of a suitable amplitude image contrast measure, regarded as a function of the reconstruction distance parameter. Here we show that, by using the sparsity measure coefficient regarded as a refocusing criterion in the holographic reconstruction, it is possible to recover the focus plane and, at the same time, establish the degree of sparsity of digital holograms, when samples of the diffraction Fresnel propagation integral are used as a sparse signal representation. We employ a sparsity measurement coefficient known as Gini's index thus showing for the first time, to the best of our knowledge, its application in DH, as an effective refocusing criterion. Demonstration is provided for different holographic configurations (i.e., lens and lensless apparatus) and for completely different objects (i.e., a thin pure phase microscopic object as an in vitro cell, and macroscopic puppets) preparation.

Enhancing depth of focus in tilted microfluidics channels by digital holography.

In this Letter we propose a method to enhance the limited depth of field (DOF) in optical imaging systems, through digital holography. The proposed approach is based on the introduction of a cubic phase plate into the diffraction integral, analogous to what occurs in white-light imaging systems. By this approach we show that it is possible to improve the DOF and to recover the extended focus image of a tilted object in a single reconstruction step. Moreover, we demonstrate the possibility of obtaining well-focused biological cells flowing into a tilted microfluidic channel.

Quantitative phase maps denoising of long holographic sequences by using SPADEDH algorithm.

We propose a denoising method for digital holography mod 2π wrapped phase map by using an adaptation of the SPArsity DEnoising of Digital Holograms (SPADEDH) algorithm. SPADEDH is a l(1) minimization algorithm able to suppress the noise components on digital holograms without any prior knowledge or estimation about the statistics of noise. We test our algorithm with either general numerical simulated wrapped phase, quantifying the performance with different efficiency parameters and comparing it with two popular denoising strategies, i.e., median and Gaussian filters, and specific experimental tests, by focusing our attention on long-sequence wrapped quantitative phase maps (QPMs) of in vitro cells, which aim to have uncorrupted QPMs. In addition, we prove that the proposed algorithm can be used as a helper for the typical local phase unwrapping algorithms.

On the holographic 3D tracking of in vitro cells characterized by a highly-morphological change.

Digital Holography (DH) in microscopic configuration is a powerful tool for the imaging of micro-objects contained into a three dimensional (3D) volume, by a single-shot image acquisition. Many studies report on the ability of DH to track particle, microorganism and cells in 3D. However, very few investigations are performed with objects that change severely their morphology during the observation period. Here we study DH as a tool for 3D tracking an osteosarcoma cell line for which extensive changes in cell morphology are associated to cell motion. Due to the great unpredictable morphological change, retrieving cell's position in 3D can become a complicated issue. We investigate and discuss in this paper how the tridimensional position can be affected by the continuous change of the cells. Moreover we propose and test some strategies to afford the problems and compare it with others approaches. Finally, results on the 3D tracking and comments are reported and illustrated.

Clear coherent imaging in turbid microfluidics by multiple holographic acquisitions.

Recently it has been demonstrated that digital holography is a powerful means allowing imaging of both amplitude and phase objects in turbid flowing media. However, in quasi-static turbid microfluidics, multiple scattering contributions through the colloids superimpose coherently to the recording device, resulting in speckle noise and hindering a clear vision of the objects. In this Letter we exploit the Brownian motion of the colloidal particles to get multiple uncorrelated holograms, and we combine them to reduce the speckle contrast. In this way we get a multi-look gain without losing image resolution.

Light-Induced Dynamic Holography.

Holographic photomobile polymers (H-PMP) are a novel class of photomobile materials in which holograms can be optically recorded. They can be used in a large variety of applications, including optical switches and color selectors. In this work, we show one of the most important properties of the photomobile film, which is the photophobicity of the unpolymerized parts of the photomobile mixture. In order to investigate this property, we recorded a transmission phase grating on an H-PMP film, and used a different experimental technique to measure the diffraction efficiency, surface tension, and mixture properties. The results allowed for a better understanding of the mechanism of the light-controlled bending observed in these compounds.

Twin-beams digital holography for 3D tracking and quantitative phase-contrast microscopy in microfluidics.

We report on a compact twin-beam interferometer that can be adopted as a flexible diagnostic tool in microfluidic platforms with twofold functionality. The novel configuration allows 3D tracking of micro-particles and, at same time, can simultaneously furnish Quantitative Phase-contrast maps of tracked micro-objects by interference microscopy, without changing the configuration. Experimental demonstration is given on for in vitro cells in a microfluidic environment.

Light induced patterning of poly(dimethylsiloxane) microstructures.

A new method for direct patterning of Poly(dimethylsiloxane) (PDMS) microstructures is developed by taking advantage of photorefractive effect in a functionalized substrate. Here we show that when a x-cut Iron doped Lithium Niobate (LN) crystal is exposed to appropriate structured laser light, a charge density pattern builds-up in the crystal and a space charge field arise that is able to induce self-patterning of the PDMS liquid film deposited on its surface via the dielectrophoretic effects. Proper heating treatment allows to achieve polymeric linking process creating a solid and stable PDMS microstructures. The self-patterned structures replicate the illuminating light pattern. We show that 1D and 2D patterning of PDMS gratings can be achieved. This new soft-lithographic approach can pave the way for realizing PDMS micro-structures with high degree of flexibility that avoids the need of moulds fabrication.

Synthesis and display of dynamic holographic 3D scenes with real-world objects.

A 3D scene is synthesized combining multiple optically recorded digital holograms of different objects. The novel idea consists of compositing moving 3D objects in a dynamic 3D scene using a process that is analogous to stop-motion video. However in this case the movie has the exciting attribute that it can be displayed and observed in 3D. We show that 3D dynamic scenes can be projected as an alternative to complicated and heavy computations needed to generate realistic-looking computer generated holograms. The key tool for creating the dynamic action is based on a new concept that consists of a spatial, adaptive transformation of digital holograms of real-world objects allowing full control in the manipulation of the object's position and size in a 3D volume with very high depth-of-focus. A pilot experiment to evaluate how viewers perceive depth in a conventional single-view display of the dynamic 3D scene has been performed.

Multi-imaging capabilities of a 2D diffraction grating in combination with digital holography.

In this Letter we report on an alternative approach to get multiple images in microscopy, exploiting the capabilities of both a lithium niobate diffraction grating and digital holographic technique. We demonstrate that multi-imaging can be achieved in a lensless configuration by using a hexagonal diffraction grating but overcoming, thanks to digital holography (DH), the many constrains imposed by the grating parameters in multi-imaging with Talbot effect or Talbot array illuminators. In fact, DH permits the numerical reconstruction of the optical field diffracted by the grating, thus obtaining in-focus multiple images in a plane different from the fractional or entire Talbot ones.

Compression of digital holograms via adaptive-sparse representation.

Efficient storage and transmission of digital holograms (DHs) requires the development of appropriate compression techniques for such a special class of images. In this Letter, we investigate a method to compress DHs using a sparse matrix representation. Using digital holography to numerically manage complex wave fields, we are able to apply an adaptive mask, based on a threshold filter, to the object wave field. From there, we store the result of this filtering by sparse representation. In this Letter, we demonstrate that using sparse representation allows for a high compression factor with minimal loss in the quality of the reconstructed image. This technique is efficient for storage and transmission of DHs.

Optical reconstruction of digital holograms recorded at 10.6 microm: route for 3D imaging at long infrared wavelengths.

We demonstrate the optical reconstruction in the visible range (0.532 microm) of digital holograms recorded at long IR wavelengths (10.6 microm) by means of a spatial light modulator. By using an integrated recording-reconstruction system, it is, in fact, feasible to achieve direct imaging of holograms acquired outside the visible range, i.e., in the IR spectrum. By choosing a Fourier recording configuration, the reconstructed image, obtained at about a 20 times shorter wavelength than the acquisition image, exhibits minor aberrations, which do not significantly affect the optical reconstruction. The high NA achievable at a long IR wavelength allows us to image large objects at reasonable distances.

Compact off-axis holographic slide microscope: design guidelines.

Holographic microscopes are emerging as suitable tools for in situ diagnostics and environmental monitoring, providing high-throughput, label-free, quantitative imaging capabilities through small and compact devices. In-line holographic microscopes can be realized at contained costs, trading off complexity in the phase retrieval process and being limited to sparse samples. Here we present a 3D printed, cost effective and field portable off-axis holographic microscope based on the concept of holographic microfluidic slide. Our scheme removes complexity from the reconstruction process, as phase retrieval is non iterative and obtainable by hologram demodulation. The configuration we introduce ensures flexibility in the definition of the optical scheme, exploitable to realize modular devices with different features. We discuss trade-offs and design rules of thumb to follow for developing DH microscopes based on the proposed solution. Using our prototype, we image flowing marine microalgae, polystyrene beads, E.coli bacteria and microplastics. We detail the effect on the performance and costs of each parameter, design, and hardware choice, guiding readers toward the realization of optimized devices that can be employed out of the lab by non-expert users for point of care testing.

Creating an extended focus image of a tilted object in Fourier digital holography.

We present a new method to numerically reconstruct images on a tilted plane by digital holography in Fourier configuration. The proposed technique is based on a quadratic deformation of spatial coordinates of the digital hologram. By this approach we demonstrate that it is possible to recover the extended focus image (EFI) of a tilted object in a single reconstruction step from the deformed hologram.

Correct self-assembling of spatial frequencies in super-resolution synthetic aperture digital holography.

Synthetic aperture enlargement is obtained, in lensless digital holography, by introducing a diffraction grating between the object and the CCD camera with the aim of getting super-resolution. We demonstrate here that the spatial frequencies are naturally self-assembled in the reconstructed image plane when the NA is increased synthetically at its maximum extent of three times. By this approach it possible to avoid the use of the grating transmission formula in the numerical reconstruction process, thus reducing significantly the noise in the final super-resolved image. Demonstrations are reported in 1D and 2D with an optical target and a biological sample, respectively.